1. Field of the Invention
The present invention relates generally to the fields of cancer therapy and gene therapy. More particularly, it concerns the use of PEA3 to prevent and treat various transformation events.
2. Description of Related Art
It is well established that a variety of cancers are caused, at least in part, by genetic abnormalities that result in either the over-expression of one or more genes, or the expression of an abnormal or mutant gene or genes. For example, in many cases, the expression of oncogenes is known to result in the development of cancer. "Oncogenes" are genetically altered genes whose mutated expression product somehow disrupts normal cellular function or control (Spandidos el al., 1989).
Most oncogenes studied to date have been found to be "activated" as the result of a mutation, often a point mutation, in the coding region of a normal cellular gene, i.e., a "proto-oncogene", that results in amino acid substitutions in the expressed protein product. This altered expression product exhibits an abnormal biological function that takes part in the neoplastic process (Travali et al., 1990). The underlying mutations can arise by various means, such as by chemical mutagenesis or ionizing radiation. A number of oncogenes and oncogene families, including ras, myc, neu, raf, erb, src, fins, jun and abl, have now been identified and characterized to varying degrees (Travali et al., 1990; Bishop, 1987).
The neu gene (also known as HER-2/neu or c-erb-2) encodes a 185-kDa transmembrane tyrosine kinase (p185.sup.neu) with homology to epidermal growth factor receptor (Hung et al., 1986; Coussens et al., 1985; Schechter et al., 1984; Sanba et al., 1985; Yamamoto et al., 1986). Enhanced expression of neu is known to be involved in many human cancers, including NSCLC and has been shown to correlate with poor patient survival in NSCLC (Kern et al., 1990; Schneider et al., 1981; Weiner et al., 1990). Cellular and animal studies have shown that an increase in neu tyrosine kinase activity increases the expression of malignant phenotypes (Muller et al., 1988; Hudziak et al., 1987; Muthuswamy et al., 1994; Yu et al., 1991; Yu et al., 1993; Hung et al., 1989; Sistonen et al., 1989; Yu et al., 1994).
The neu oncogene, was first identified in transfection studies in which NIH 3T3 cells were transfected with DNA from chemically induced rat neuroglioblastomas (Shih et al., 1981). The p185 protein encoded by neu has an extracellular, transmembrane, and intracellular domain, and therefore has a structure consistent with that of a growth factor receptor (Schechter et al., 1984). The human neu gene was first isolated due to its homology with v-erbB and EGF-r probes (Senba et al., 1985).
The neu oncogene plays an important role in carcinogenesis, for example, the gene is amplified in approximately 30% of primary breast cancer. Amplified expressions of the neu oncogene in transfected 3T3 cells induces malignant transformation. neu expression has also been detected in ovarian cancer and its overexpression results in poor prognosis. The expression of neu oncogenes in human tumor cells induce resistance to several host cytotoxic mechanisms.
Along with an increased proliferative potential, neu-mediated cancers appear to be resistant to host defense mechanisms. Studies have shown that overexpression of the neu oncogene in transfected cells results in resistance to tumor necrosis factor, a major effector molecule in macrophage-mediated tumor cell cytotoxicity.
Therefore, neu oncogene expression is correlated with the incidence of cancers of the human breast and female genital tract. Moreover, amplification/overexpression of this gene has been directly correlated with relapse and survival in human breast cancer (Slamon et al., 1987; 1989). Therefore, it is important to evolve information regarding the neu oncogene, particularly information that could be applied to reversing or suppressing the oncogenic progression that seems to be elicited by the presence or activation of this gene. Unfortunately, little has been previously known about the manner in which one may proceed to suppress the oncogenic phenotype associated with the presence of oncogenes such as the neu oncogene.
In addition, neu overexpression in NSCLC is associated with shortened survival. In vitro experimental models have provided evidence that, in the murine cell NIH 3T3, oncogenes increase drug resistance. Furthermore, Tsai et al., 1993 and 1995 used a NSCLC model to demonstrate that activation of an oncogene is quantitatively associated with intrinsic chemoresistance in human malignant cells. This resistance is observed with a variety of drugs that are structurally unrelated and act on different targets and/or by different mechanisms. Thus increased expression of neu oncogene enhances chemoresistance to a wide variety of chemotherapeutic agents (Tsai, 1993) including cisplatin, doxorubicin, and VP16 (Tsai et al., 1993; Tsai et al., 1995). The association of neu overexpression in cancer cells with malignant phenotypes and chemoresistance provides a plausible interpretation for the poor clinical outcome for patients with neu overexpressing tumors.
Although breast cancer diagnosed in its earliest clinical stages (stage 0, stage Ia) is highly curable, the cure rate for more advanced stages drops precipitously, even after modem combined-modality treatments. Metastatic breast cancer responds to both chemotherapy and hormone therapy, and most patients can be palliated adequately during the 1 to 3 years of usual survival. However, metastatic breast cancer is considered incurable, as demonstrated by the relentless death rates, regardless of the treatment modality utilized. Front-line chemotherapy or hormone therapy programs for correctly selected patients produce objective responses in 50% to 70% of patients, but the median duration of response is usually less than one year. Response rates after second line treatments are considerably lower (20% to 50%), and response durations average 6 months.
Ovarian cancer is also highly curable in its earliest stages, but the overwhelming majority of patients are diagnosed in stages III and IV. Although responsive to chemotherapy, most patients with advanced ovarian cancer relapse and die of their disease. With the introduction of several neu cytotoxic agents (taxanes, vinorelbine, platinum derivatives), some responses are observed after second line therapy too, but cure in this situation remains an elusive goal.
Thus overexpression of the HER-2/neu oncogene correlates with poor survival for breast and ovarian cancer patients and induces metastatic potential and chemoresistance of human cancer cells. Repression of HER-2/neu suppresses the malignant phenotypes of HER-2/neu-overexpressing cancer cells. It suggests that HER-2/neu oncogene is an excellent target for development of novel therapeutic agent for the HER-2/neu-overexpressing cancer cells. Thus methods and compositions that repress HER-2/neu transcription in HER-2/neu-overexpressing human breast and ovarian cancer cell lines, and suppress activated neu induced transformation would be of great therapeutic value in the treatment of these diseases. PEA3, as a transcription factor, targets HER-2/neu gene by repressing its expression, thus it has a great potential to be used as a therapeutic strategy of these neu-mediated cancer types.
In 10-20% of the HER2 overexpressing breast tumors, some gastric and virtually all HER2.sup.+ lung cancers HER2 mRNA and protein overexpression occur in the absence of increased gene copy number thus suggesting that HER2 there may be some aberration in transcriptional regulation that plays a fundamental role in the development of these diseased states.
It has been shown that neu transcription can be enhanced by a variety of growth regulatory agents such as phorbol esters, epidermal growth factor and dibutyryl cAMP. Studies with neu promoters have identified cis and trans acting elements that may be involved in the regulation of neu transcription. Many DNA-binding trans-acting proteins are capable of stimulating DNA replication as well as gene transcription. The identification of a specific neu transactivator potentially leads to a molecular understanding of the development of neu gene amplification.
Recent studies showed that there is an ETS response element that is conserved within a Dnase I hypersensitive site in the proximal HER2 promoter region. This study concluded that ETS factors direct the overexpression of many gene products critical for human breast tumorogenesis. In yet another study, it was demonstrated that PEA3, a newly identified member of the ETS family is over-expressed in mouse metastatic mammary adenocarcinoma.
In other, contradictory studies when ETS-1 was ectopically expressed in two different highly tumorogenic human colon cancer cell lines it reversed the transformed phenotype and tumorogenicity in a dose dependent manner (Suzuki et al., 1995). In yet another study raised the question of suppresser activity for some ETS-1 products in T-cell acute lymphoblastic leukemias.
Hence it appears that there is much confusion regarding the putative role of the ETS family of transcription regulators.